1. Field of the Invention
The present invention relates to new compounds having high affinity and selectivity for the melanocortin 1 receptor (MC1-receptor). The new compounds selectively activate or block the MC1-receptors, which makes it possible to discriminate between the physiological effects mediated by the MC1-receptors and other subtypes of melanocortin receptors, and to afford selective pharmacological actions upon their administration to an animal or a human. The invention also relates to methods for their manufacture and their pharmaceutical preparations, as well as to their use for the treatment of medical and veterinary conditions which can be influenced by the MC1-receptor.
2. Related Art
Melanocortic peptides (melanocortins) are natural peptide hormones of animals, in particular mammals including man, which bind to and stimulate MC-receptors. Examples of melanocortins are α-MSH, β-MSH, γ-MSH, ACTH, and their peptide fragments. α-MSH is mainly known for its ability to regulate peripheral pigmentation (Eberle 1988), whereas ACTH is known to induce steroidoneogenesis (Simpson and Waterman 1988). The melanocortic peptides also mediate a number of other physiological conditions. Thus, they are reported to act as immunomodulators and to affect one or more of motivation, learning, memory, behaviour, inflammation, body temperature, pain, perception, blood pressure, heart rate, vascular tone, brain blood flow, nerve growth, placental development, aldosteron synthesis and release, thyroxin release, spermatogenesis, ovarian weight, prolactin and FSH secretion, uterine bleeding in women, sebum and pheromone secretion, blood glucose levels, weight homeostasis, intrauterine fetal growth as well as other events surrounding parturition (Garrud et al., 1974, Wiegant et al., 1979, O'Donahue et al., 1981, O'Donahue & Dorsa 1982, DeWied & Jolles 1982, Klein et al., 1985, Feng et al. 1987, Lin et al. 1987, Eberle, 1988, Gruber & Callahan 1989, DeWildt et al., 1995. Fruedman 1997).
By the use of molecular cloning, genes encoding five different subtypes of MC-receptors have been identified (Chhajlani et al. 1993, Chhajlani and Wikberg 1992, Gantz et al. 1993a, b, Mountjoy et al. 1992). The MC-receptors belong to the class of G-protein coupled receptors which are all built from a single peptide forming 7 transmembrane domains. The five MC-receptors are termed MC1, MC2, MC3, MG4 and MC5 and they all couple in a stimulatory fashion to cAMP. Of these the MC2-receptor is the ACTH-receptor whereas the others constitute subtypes of melanocyte stimulating hormone receptors (MSH-receptors).
The MC1-receptor is present on melanocytes and melanoma cells (Low et al. 1994, Siegrist & Eberle 1995). Recent data also indicate that the MC1-receptor is expressed in limited areas (periaqueductal gray) of rat and human brain (Xia et al. 1995), as well as in the testis (Vanetti et al. 1994). Also, very importantly recently the MC1-receptor is shown to be present on macrophages (Star et al. 1995), neutrophils (Catania et al. 1996), glioma cells and astrocytes (Wong et al. 1997), monocytes and endothelial cells (Hartmeyer et al. 1997, and references therein). Recently the MC1-receptor mRNA was also detected in the normal mouse brain (Rajora et al. 1997a), suggesting a specific function of this receptor in the brain. Moreover, recent data obtained by immunohistochemical detection techniques show that MC1-receptors are present in testis and ovary (Thömvall et al. 1997). This suggests a specific role of the MC1-receptor in reproductive physiology, as well as possibly in reproductive pathophysiology.
The MC2-receptor is the ACTH receptor. It is present in the cortex of the adrenal gland. The MC3-receptor mRNA is found in distinct areas of the brain, as well as in placental and gut tissues (Gantz et al. 1993a, Desarnaud et al. 1994, Roselli-Rehfuss et al. 1993). The MC4-receptor is ubiquitous in the brain (Gantz et al. 1993b, Mountjoy et al, 1994). The MC5-receptor is expressed in the brain, as well as in several peripheral tissues (Chhajlani et al. 1993, Gantz et al. 1994, Griffon et al. 1994, Labbé et al. 1994, Barrett et al. 1994, Fathi et al. 1995). More recent data indicate that all the 5 cloned MC-receptors have a wider tissue distribution (Chhajlani 1996).
The five MC-receptors show unique affinities for the melanocortic peptides (Schiöth et al. 1995, Schiöth et al. 1996a,b,c). Thus, the MC1-receptor shows high affinity for α-MSH, but lower affinities for β-MSH, γ-MSH and ACTH. The MC2-receptor binds ACTH with high affinity, but it does not bind the MSH peptides. The MC3-receptor show slightly higher affinity for γ-MSH compared to β- and α-MSH. The MC4-receptor shows slight preference for β-MSH, over α-MSH, and a very low affinity for γ-MSH. The MC5-receptor shows the same potency order for the MSH peptides as the MC1-receptor, although with much lower affinities (Schiöth et al. 1995, Schiöth et al. 1996a,b,c).
A diversity of effects is induced by natural melanocortic peptides not yet fully related to the various MC-receptor subtypes. These effects should be mediated by different subtypes of the MC-receptors. Most pertinent, however, is that increasing evidence indicates the MC1-receptors play an important role in the modulation of inflammation. Thus, for example, α-MSH was shown to inhibit formation of nitric oxide (NO) in cultured murine macrophages stimulated with bacterial lipopolysaccharide and γ-interferon, an effect claimed to be caused by the inhibition of the production of NO synthase (NOS) by the stimulation of MC1-receptors in macrophages (Star et al. 1995). As NO is believed to be a common mediator of all forms of inflammation this indicates that stimulation of MC1-receptors mediates the anti-inflammatory effect earlier known to be induced by MSH-peptides. α-MSH is also known to increase the formation of interleukin 10 (IL-10) in monocytes, which is believed to be an important component in immunosuppressive effects induced by α-MSH (Bhardwaj et al. 1996).
Recent studies also show that α-MSH potently inhibits the chemotactic migration of neutrophils (Catania et al. 1996). Moreover, neutrophils were shown to contain MC1-receptor mRNA, which was upregulated on stimulation of the neutrophils with interferon and bacterial lipopolysaccharide (Catania et al. 1996). Thus, as neutrophil migration constitutes an important component in early inflammation, these results again indicate the importance of the MC1-receptor as mediators of inhibition of inflammation.
In another study the injection of α-MSH, as well as the MSH-analogue [Nle4,D-Phe7]α-MSH (NDP-MSH) was shown to inhibit the release of cytokines IL-1 and TNF-α into the blood after intra-peritoneal injection of lipopolysaccharide (Goninard et al. 1996). This supports the anti-inflammatory role of MSH-peptides.
Important anti-inflammatory roles of MC-receptors (presumed to be of the MC1 type) have also been implicated in the brain since α-MSH inhibits the production of tumour necrosis factor alpha (TNF-α) in vivo, as well as in vitro on glioma cells; in the later case α-MSH was shown to inhibit formation of TNF-α induced by bacterial endotoxin (Wong et al. 1997). In another study α-MSH given intracerebroventricularly or intraperitonally inhibited formation of central TNF-α induced by locally administered bacterial lipopolysaccharide (Rajora et al. 1997a). TNF-α occurs in neurological disorders, infection and injury of the brain, and is thought to underlie pathological processes in the brain. These data indicate an important role of MC-receptors as mediators of central anti-inflammatory actions.
Recently α-MSH was also shown to reduce inflammation in a model for inflammatory bowel disease (Rajora et al. 1997b).
The α-MSH peptide too is ascribed an important role in cutaneous biology. Most well known is its ability to stimulate pigment formation of the skin. α-MSH may act not only on MC-receptors located to melanocytes but also on immunocompetent and inflammatory cells, keratinocytes, fibroblasts and endothelial cells of the skin, thereby modifying keratinocyte proliferation and differentiation, and regulate endothelial cell and fibroblast cytokine production, as well as fibroblast collagenase production. α-MSH is known to down-regulate the production of proinflammatory cytokines and accessory molecules on antigen presenting cells. In contrast suppressor factors such as IL-10 are upregulated by α-MSH (Luger 1997). In vivo data show that systemic application of α-MSH inhibits the induction and elicitation of contact hypersensitivity and induces hapten tolerance (Luger 1997). Thus, the accumulating evidence indicates that the stimulation of MC-receptors, presumably of the MC1-receptor subtype, mediates important negative regulation mechanisms of cutaneous inflammation and hyper-proliferative skin diseases (Luger 1997).
In addition to these findings, Hartmeyer et al. (1997) have shown that α-MSH increases MC1-receptor expression in dermal microvasculature endothelial cells and causes increased release of interleukin 8 (IL-8) from these cells. This indicates a role of MC1-receptors in the skin as modulators of inflammation and immunity (see Hartmeyer et al. 1997).
For further reading on the anti-inflammatory role of MSH peptides reference is made to the review by Lipton and Catania (1997).
Since 1957 MSH-receptors have been known as physiological entities. Binding sites for MSH/ACTH peptides have been identified in number of brain and peripheral tissues (Hnatowich et al. 1989, Tatro & Reichlin 1987, Lichtensteiger et al. 1993, Tatro & Entwistle 1994). Peptide structure activity studies of these receptors have been performed on melanophores from lower vertebrates like Rana pipiens (frog), Anolis carolinensis (lizard) and Xenopus laevis. Receptor studies were later also performed by binding on melanoma cell lines. These test systems gave comparable results and it is now known that the data obtained with these systems refer to the MC1-receptor (Eberle 1988).
Using such test systems it was found that replacement of L-Phe by D-Phe in α-MSH provided high potency and prolonged action (Sawyer et al., 1980). Cyclic [Cys4, Cys10]α-MSH analogues were also synthesized; they were found to be potent melanotrophs in skin pigmentation bioassays (Knittle et al., 1983, Sawyer et al., 1982). However, while some of the previous known natural and synthetic peptides, as well as more recently synthesized ones, show high affinity for MC1-receptors, their selectivity versus other subtypes of MC-receptors are limited (see e.g. Hol et al. 1994, Adan et al. 1994, among others). Recently, however, a peptide found by phage display screening was described which showed higher selectivity for MC1-receptors compared to other subtypes for MC receptors (Szardenings et al. 1997). However, for clinical and other uses this peptide showed inferior properties due to 1) comparatively low affinity for MC1-receptors, 2) low agonistic effects on MC1-receptors compared to other peptide hormones and 3) low stability due to presence of oxidizable SH groups and instability in regard of proteolytic cleavage.
There remains a need to provide means and methods to selectively regulate MC1-receptors. Thereby pharmacological effects affecting processes and conditions related to tissues and cells expressing the MC1-receptor may be elicited. These processes and conditions comprise but are not limited to immune responses, inflammatory processes, imunotolerance, immunomodulation, allergic processes, reproductive processes, melanoma and malignant diseases related to MC1-receptor expressing cells.
There is furthermore a need to provide chemical compounds that activate MC1-receptors selectively and with high potency; to provide chemical compounds which antagonize the action of other hormones and agonists on MC1-receptors selectively and with high potency; and to provide a method for administration of said compounds to animals including man.